Background: Tooth enamel is the hardest and most highly mineralized tissue in the human body. It serves as a protective barrier against chemical, mechanical, and microbial challenges. Despite its durability, enamel remains vulnerable to developmental and posteruptive defects such as fluorosis, hypomineralization, and amelogenesis imperfecta (AI). For oral health professionals, a clear understanding of the biological and molecular mechanisms underlying enamel formation is essential for advancing preventive and therapeutic strategies in clinical practice. This review synthesizes current knowledge on enamel formation, with emphasis on its cellular, molecular, and structural determinants, and discusses clinically relevant disruptions as well as emerging biomimetic approaches. Methods: This scoping review was conducted according to the PRISMA-ScR guidelines. A systematic literature search of the mechanisms of enamel formation was performed via Embase and Medline. Titles and abstracts were screened independently by three authors. Studies that primarily addressed enamel defects were excluded from the systematic synthesis; however, these studies were retained for narrative discussion. Following the screening process, 92 publications met the inclusion criteria and were incorporated into the thematic synthesis. Results: Enamel formation is a complex, multistage process involving epithelial-mesenchymal interactions and the sequential activity of ameloblasts during presecretory, secretory, transition, and maturation stages. Key mechanisms include the secretion of enamel matrix proteins (e.g., amelogenin, ameloblastin, and enamelin), proteolytic processing by enzymes such as MMP20 and KLK4, and controlled ion transport, leading to hydroxyapatite crystal growth and organization into rod and interrod structures. The structural arrangement endows enamel with exceptional mechanical resistance. Narrative sections address "What can go wrong?", summarizing genetic, epigenetic, and environmental causes of fluorosis, hypomineralization, and amelogenesis imperfecta, and other developmental defects, whereas "What can we learn from nature?" highlights biomimetic strategies. Conclusions: Human enamel formation is a highly coordinated biomineralization process regulated at the cellular, structural, and molecular levels. Disruptions in these processes underlie major enamel pathologies. Integrating mechanistic insights from natural enamel development with emerging biomimetic technologies offers promising avenues for prevention, diagnosis, and treatment in dentistry. This review provides oral health professionals with a biologically grounded framework to guide evidence-based management of enamel-related conditions.

This study aims to investigate the qualitative and quantitative morphological changes in dental enamel resulting from SHS exposure during early childhood, utilizing an animal model. Wistar rat offspring were divided into a control group (n = 15) and an SHS exposure group (n = 19). The experimental group was subjected to cigarette smoke twice daily for 5 min, from postnatal day 2 to day 28. Throughout the experimental period, growth was monitored by measuring both weight and body length. Dental enamel was assessed through photographic analysis, micro-computed tomography (micro-CT), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDS), and microhardness testing. Mandibles were processed, followed by hematoxylin and eosin staining. Data analysis was conducted using Student’s t-tests (⍺= 5

PURPOSE:The junctional epithelium (JE) covers the cervical areas of developing or existing teeth. It can re-establish itself even after being removed during periodontal therapies, followed by wound healing. However, the mechanisms that can maintain this universally conserved structure are still unclear. METHODS:The molecular mechanisms of JE homeostasis were investigated by altering levels of JE-specific genes in a telomerase immortalized human gingival epithelial cell line (TIGKs) by exposing TIGKs to conditioned medium (C-CM) from cultivated human cementoblasts. The mRNA and protein levels of JE-associated genes in TIGKs were examined using real-time polymerase chain reaction (PCR) and immunocytochemistry (ICC) after treatment with C-CM. The candidate pathways related to differential mRNA and protein expression were analyzed with a DNA microarray and identified using Kyoto Encyclopedia of Genes and Genomes and WikiPathways. Real-time PCR and ICC were conducted to confirm the changes in the expressions of candidate genes. RESULTS:mRNA levels and protein expressions of amelotin (Amtn) were upregulated after treatment with C-CM for 48 hours. DNA microarray analyses identified 595 genes that were upregulated >2-fold, and 820 genes that were downregulated >2-fold. C-CM promoted the expression of suppressor of cytokine signaling 3 and reduced the expression of an inactivator of Janus kinase 2. Both signaling molecules were found, using siRNA technology, to mediate the increase of Amtn mRNA and protein expression levels. CONCLUSIONS:The upregulation of Amtn in gingival epithelial cells by C-CM suggests that this regulatory pathway is associated with the homeostasis of JE structures by the cementum.

Objective The junctional epithelium (JE) is composed of tightly interconnected layers of squamous epithelial cells that play a crucial role as a defence mechanism. Recently, several enamel-derived proteins have been shown to play a role in the adhesion of JE cells to the mineralised tooth surface. This study aims to explore the in vivo protein expression levels of amelotin (AMTN), laminin (LAM332) and the protein secreted by follicular dendritic cells (FDC-SP) within the JE, both in the presence and absence of experimental periodontitis (EP) in rats.Materials and methods In total, 16 rats were randomly divided into two groups: a control group without EP and a group with induced periodontitis. EP was established by placing cotton ligatures around the cervical region of the lower first molars. Fifteen days after EP induction, all animals were euthanised and mandibles were harvested. Micro-computed tomography, histomorphometric, and immunohistochemistry analyses were employed to assess volumetric bone alterations, architectural bone parameters and number of osteoclasts. The presence of inflammatory cells and enamel protein expression were evaluated by immunofluorescence.Results The results demonstrated that the EP group showed a significant increase in alveolar bone loss, an elevated number of tartrate-resistant acid phosphatase-positive cells, and enhanced inflammatory processes compared to the control group. Immunofluorescence staining revealed a significant increase in the expression of AMTN in the EP group. However, there were no significant differences between the expression of FDC-SP and LAM332. Correlation analysis of AMTN, LAM332 and FDC-SP with the intensity of inflammatory markers (CD45+, CD66b+ and CD8+, CD163+ and CD80+) showed no significant differences for the EP group.Conclusion Our data suggest that the expression of AMTN increased after inflammatory stimuli and that AMTN may be associated with the onset and progression of EP.

Achieving superior mechanical properties of composite materials in artificially engineered materials is a great challenge due to technical bottlenecks in the size and morphological modulation of inorganic nanominerals. Hence, a "bioprocess-inspired fabrication" is proposed to create multilayered organic-inorganic columnar structures. The sequential assembly of halloysite nanotubes (HNTs), polyelectrolytes (PAAs), and calcium phosphates (CaPs) results in organic-inorganic structures. PAA plays a crucial role in controlling the formation of CaP, guiding it into amorphous particles with smaller nanosizes. The introduction of HNT induces the assembly and maturation of CaP-PAA, leading to the formation of a highly crystalline hydroxyapatite. Poly(vinyl alcohol) was then woven into HNT-encapsulated hydroxyapatite nanorods, resulting in composite materials with basic hierarchical structures across multiple scales. The fabricated composite exhibits exceptional hardness (4.27 +/- 0.33 GPa) and flexural strength (101.25 +/- 1.72 MPa), surpassing those of most previously developed biological hard tissue materials. Additionally, the composite demonstrates effective antibacterial properties and corrosion resistance, attributed to the dense crystalline phase of CaP. This innovative approach showcases the potential of clay minerals, particularly HNT, in the advancement of biomaterial design. The outstanding mechanical and antimicrobial properties of clay-based composites make them a promising candidate for applications in hard tissue repair, offering versatility in biomedicine and engineering.

Abstract Background This study aimed to evaluate dentin wear and biological performance of desensitizing materials. Methods Seventy bovine root dentin blocks were sectioned. Half of the surface of each specimen was untreated (control) and the other half was immersed in EDTA and treated with the following desensitizing materials: placebo varnish (PLA), fluoride varnish (FLU), sodium fluoride (NaF) varnish + sodium trimetaphosphate (TMP), universal adhesive (SBU), S-PRG varnish (SPRG), biosilicate (BIOS), and amelotin solution (AMTN). After application, the specimens were submitted to an erosive-abrasive challenge and the wear analyzed by optical profilometer. Serial dilutions of extracts obtained from the culture medium containing discs impregnated with those desensitizers were applied on fibroblasts and odontoblasts-like cells cultures. Cytotoxicity and production of total protein (TP) by colorimetric assays were determined after 24 h. Data were statistically analyzed using Kruskal-Wallis, Dunn’s, One-way ANOVA and Tukey tests (p ≤ 0.05). Results No dentin wear was observed only for SBU. The lowest dentin wear was observed for AMTN and TMP. Cell viability was significantly reduced after treatment with undiluted extracts of PLA, FLU, TMP and SBU in fibroblasts and TMP and SBU in odontoblast-like cells. SPRG, BIOS and AMTN were cytocompatible at all dilutions tested. Considering TP results, no statistical difference was observed among the groups and high levels for TP were observed after TMP and FLU treatments. Conclusions Universal adhesive system may protect dentin with opened tubules from wear after challenge. Extracts of adhesive and fluoride varnishes presented cytotoxic mainly on fibroblasts. The enamel protein may be a future alternative to treat dentin with opened tubules because it may cause low wear under erosive-abrasive challenge with low cytotoxic effects.

Recent years have seen significant positive changes and developments in oral health-related policy and data on oral health and oral health care in Canada. Simultaneously, on the international stage, the momentum for oral health and related research continues to build. These changes have led to an initiative to create Canada's first National Oral Health Research Strategy (NOHRS), which was recently published by the Canadian Institutes of Health Research-Institute of Musculoskeletal Health and Arthritis (Allison and Rock 2024). In this communication, we describe the process that was used to undertake this work. We present the resulting guiding principles, the research priority areas, and the framework that emerged, which included 6 strategic priorities grouped into 3 themes: (A) Leading Issues: (1) access to care, (2) inequities, identities, and oral health; (B) Emerging Methods: (3) artificial intelligence, (4) omics; and (C) Overarching Approaches: (5) environmental sustainability, (6) knowledge mobilization and implementation science. In addition, NOHRS includes a series of proposed goals and a timeline over the coming years. The point is to encourage a broad range of individuals and groups of people to engage with this high-level strategy and create plans to implement it. This strategy directly answers the call by the World Health Organization for countries to establish a national oral health research strategy (World Health Organization 2024). We have engaged in an extensive, broad consultative process, resulting in a Canadian NOHRS that is tailored to the needs of our community. Its aim is to galvanize our community into action to address the priorities we have identified. By engaging in this process, we build upon multiple oral health-related initiatives in Canada and on the international stage. We hope to inspire and facilitate similar, much-needed work elsewhere.

Abstract Background White spot lesions (WSL) represent the earliest stage of caries formation in which mineral is lost from the enamel surface, but the surface retains its integrity. At this stage, remineralization of enamel is generally considered possible. This study aimed to develop a reliable in vitro protocol for the creation of artificially induced WSL and to examine the WSL by micro-computed tomography (microCT) and optical coherence tomography (OCT). Methods Artificial WSL lesions were created by immersing human molars in a lactic acid solution under constant agitation at 37ºC for seven days. MicroCT and OCT were used to image the lesions before comparing them to naturally occurring WSL. In addition, the mineral density of the demineralized enamel and the depth of the lesion was characterized directly on the acquired images. Results The average mineral density of artificial WSL was 1.57 ± 0.21 g/cm3, compared to sound enamel with a mean mineral density of 2.9 ± 0.06 g/cm3. The mean lesion depth of 167.76 ± 0.03 µm for artificial WSL varied slightly between individual samples. The artificial WSL did have a highly mineralized surface overlying the body of the lesion, which is characteristic of subsurface lesions; however, the lesion itself was shallower when compared to naturally occurring WSL. The OCT also detected WSL and provided an estimate of lesion depth and distance from Conclusion In summary, we have developed an experimental in vitro protocol to create artificial WSL that mimics natural caries lesions. OCT produced live scans, which allowed the detection of WSL, whereas the microCT measurements provided precise information on lesion depth and mineral density.

BackgroundThe dental pellicle is a thin layer of up to several hundred nm in thickness, covering the tooth surface. It is known to protect the teeth from acid attacks through its selective permeability and it is involved in the remineralization process of the teeth. It functions also as binding site and source of nutrients for bacteria and conditioning biofilm (foundation) for dental plaque formation.MethodsFor this updated literature review, the PubMed database was searched for the dental pellicle and its composition.ResultsThe dental pellicle has been analyzed in the past years with various state-of-the art analytic techniques such as high-resolution microscopic techniques (e.g., scanning electron microscopy, atomic force microscopy), spectrophotometry, mass spectrometry, affinity chromatography, enzyme-linked immunosorbent assays (ELISA), and blotting-techniques (e.g., western blot). It consists of several different amino acids, proteins, and proteolytic protein fragments. Some studies also investigated other compounds of the pellicle, mainly fatty acids, and carbohydrates.ConclusionsThe dental pellicle is composed mainly of different proteins, but also fatty acids, and carbohydrates. Analysis with state-of-the-art analytical techniques have uncovered mainly acidic proline-rich proteins, amylase, cystatin, immunoglobulins, lysozyme, and mucins as main proteins of the dental pellicle. The pellicle has protective properties for the teeth. Further research is necessary to gain more knowledge about the role of the pellicle in the tooth remineralization process.

Abstract Background White spot lesions (WSL) represent the earliest stage of caries formation in which mineral is lost from the enamel surface, but the surface retains its integrity. At this stage, remineralization of enamel is generally considered possible. This study aimed to develop a reliable in vitro protocol for the creation of artificially induced WSL and to examine the WSL by micro-computed tomography (microCT) and optical coherence tomography (OCT). Methods Artificial WSL lesions were created by immersing human molars in a lactic acid solution under constant agitation at 37ºC for seven days. MicroCT and OCT were used to image the lesions before comparing them to naturally occurring WSL. In addition, the mineral density of the demineralized enamel and the depth of the lesion was characterized directly on the acquired images. Results The average mineral density of artificial WSL was 1.57 ± 0.21 g/cm 3 , compared to sound enamel with a mean mineral density of 2.9 ± 0.06 g/cm 3. The mean lesion depth of 167.76 ± 0.03 µm for artificial WSL varied slightly between individual samples. The artificial WSL did have a highly mineralized surface overlying the body of the lesion, which is characteristic of subsurface lesions; however, the lesion itself was shallower when compared to naturally occurring WSL. The OCT also detected WSL and provided an estimate of lesion depth and distance from Conclusion In summary, we have developed an experimental in vitro protocol to create artificial WSL that mimics natural caries lesions. OCT produced live scans, which allowed the detection of WSL, whereas the microCT measurements provided precise information on lesion depth and mineral density.

The dento-gingival junction comprises multiple epithelia including the junctional epithelium (JE), which is the most coronally-located structural element of the dento-gingival junction that demarcates external from internal periodontal environments. After tooth eruption into the oral cavity, a specialized basal lamina is formed that provides a firm attachment of the JE to the enamel. This attachment prevents microbial species and oral debris from entering subjacent periodontal tissues. Here we discuss the expression of certain JE adhesion molecules and enamel proteins that maintain the health of the dento-gingival junction but that are perturbed in the pathogenesis of periodontitis. We also consider how evolutionary processes have influenced the development of the JE as a specialized adhesion that is well-suited for protection of the dento-gingival junction. A detailed understanding of the biology of the JE will deepen current models of dento-gingival adhesion, potentially clarify inter-patient variability of susceptibility to periodontitis and help to identify new roles of enamel proteins in periodontal regeneration.

Hydroxyapatite, Ca 5 (PO 4 ) 3 (OH), is a bioinspired active ingredient for preventive oral health care. The use of hydroxyapatite in the prevention of oral diseases has gained interest, particularly in recent years. Hydroxyapatite can be used in office and for daily oral care (e.g. in toothpastes and mouthwashes). The first clinical efficacy studies on caries prevention and reduction of symptoms of dentin hypersensitivity were conducted in the 1980s. These were followed by various in vitro and in situ studies and several more recent clinical trials. A number of systematic reviews and meta-analyses on the use of hydroxyapatite in preventing oral health problems have been published. Summarizing these data, hydroxyapatite is a versatile active ingredient with multiple benefits that include caries prevention, relief from dentin hypersensitivity and tooth whitening. Since this calcium phosphate mineral has an excellent biocompatibility, it is safe for all patient groups. This interdisciplinary review gives an overview of the developments in hydroxyapatite research, highlights the research progress made regarding hydroxyapatite as a biomimetic active ingredient and summarizes the state-of-the-art evidence in support of hydroxyapatite efficacy.

OBJECTIVES Current methods for periodontal regeneration do not promote collagen fiber insertions into new bone and cementum. We used a pig wound model to screen different functionalized collagen membranes in promoting periodontal reattachment to root surfaces. METHODS Treatment groups included (1) control with no membranes, (2) collagen-coated membranes, (3) membranes with insulin-like growth factor-1 (IGF-1), (4) membranes with amelotin, or (5) membranes attached with calcium phosphate cement (CPC), or with CPC combined with IGF-1. Flap procedures were performed on mandibular and maxillary premolars of each pig. RESULTS Histomorphometric, micro-CT, and clinical measurements obtained at 4 and 12 weeks after surgery showed cementum formation on denuded roots and reformation of alveolar bone, indicating that the pig model can model healing responses in periodontal regeneration. Calcium phosphate cement simplified procedures by eliminating the need for sutures and improved regeneration of alveolar bone (p < 0.05) compared with other treatments. There was a reduction (p < 0.05) of PD only for the IGF group. Large observed variances between treatment groups indicated that a priori power analyses should be conducted to optimize statistical analysis. CONCLUSIONS Pigs can model discrete elements of periodontal healing using collagen-based, functionalized membranes. Screening indicates that membrane anchorage with calcium phosphate cements improve regeneration of alveolar bone.

Children are prone to develop dental caries. This is supported by epidemiological data confirming early childhood caries (ECC) as a highly prevalent disease affecting more than every second child worldwide. ECC is known to result from an imbalance between re- and demineralization where demineralization dominates due to frequent acid production by cariogenic bacteria present in oral biofilms. The application of oral care formulations containing remineralizing agents helps to prevent dental caries. As young children are sensitive and usually swallow (intended or unintended) a majority of toothpaste or other oral care products during daily dental care, all ingredients, especially the actives, should be non-toxic. Biomimetic hydroxyapatite [HAP; Ca5(PO4)3(OH)] is known to have favorable remineralizing properties combined with an excellent biocompatibility, i.e., it is safe if accidently swallowed. Several clinical trials as well as in situ and in vitro studies have shown that HAP remineralizes enamel and dentin. Remineralization occurs due to deposition of HAP particles on tooth surfaces forming mineral-mineral bridges with enamel crystals, but also indirectly through calcium and phosphate ions release as well as HAP's buffering properties in acidic environments (i.e., in plaque). HAP induces a homogenous remineralization throughout the subsurface enamel lesions. This review summarizes the current evidence showing HAP as an effective remineralizing agent in oral care products for children. Additional studies showing also further beneficial effects of HAP such as the reduction of biofilm formation and the relief of hypersensitivity in children with molar incisor hypomineralization (MIH). It can be concluded that HAP is an effective and safe remineralizing agent for child dental care.

This study investigates the dentin permeability (by hydraulic conductance) and tubule occlusion (by confocal and scanning electron microscopies) of in-office desensitizing materials. Bovine dentin blocks were immersed in EDTA to open dentinal tubules. Placebo varnish (PLA), fluoride varnish (FLU), NaF 5

Interfacial failure at the resin-dentin interface is a significant disadvantage of resin-based dental restoration. In this study, we created bio-inspired bio-nano complexes using the enamel protein amelotin (AMTN) or AMTN with an engineered collagen-binding site (AMTN-Col) to coat hydroxyapatite nanoparticles (HANP). The resulting nano-bio complexes, AMTN-HANP and AMTN-Col-HANP, were evaluated for their ability to promote collagen mineralization. Our study comprises three separate phases. In phase I, developing a method for functionalizing HANP with AMTN/AMTN-Col was explored. HANP were synthesized and characterized using TEM, SAED-TEM, XRD and ATR-FTIR. The nanoparticles were functionalized with AMTN or AMTN-Col. The successful coating of the nanoparticles with the proteins was confirmed using a TEM image of immunogold-labelled samples. In phase II of the study, the mineralization potential of the synthesized bio-nano complexes was studied using model systems consisting of simulated body fluid (SBF), polymerized collagen gels, and dentin disks prepared from human extracted molars. Mineral formation in SBF was recorded with a light scattering assay using a microplate reader on 8 replicates of each sample per study time point. Statistical analysis was performed using one-way ANOVA and the Tukey test. Significance was assigned at P < 0.01. The extent of mineral formation on collagen gel and remineralization of demineralized dentin was studied with SEM. Accelerated mineral formation collagen mineralization of bio-nano complexes treated samples were observed in all model systems. In phase III of the study, the clinical utilization of AMTN/AMTN-Col coated HANP in bio-integration and enhancing the bond strength of a resin-based dental restoration and the dentin interface was investigated. The bio-nano complexes were applied as a pretreatment on dentin disks prepared from human extracted molars prior to the composite resin restoration. The micro-shear bond strength test was done on 8 samples per treatment group (a total of 32 samples). Statistical analysis on shear bond strength was performed using one-way ANOVA and the Tukey test. Significance was assigned at P < 0.01. Shear bond strength values indicated that pretreatment of dentin with the bio-nano complexes before adhesive application significantly improved shear bond strength. Conclusion: We have shown that AMTN based bio-nano complexes promote mineral formation on collagenous interfaces. Our findings can be the basis of new bio-inspired, bio-nano materials that may improve dental restoration longevity by enhancing the stability and integrity of the dentin-composite resin interface.

Common periodontal disease treatment procedures often fail to restore the structural integrity of the junctional epithelium (JE), the epithelial attachment of the gum to the tooth, leaving the tooth-gum interface prone to bacterial colonization. To address this issue, we introduced a novel bio-inspired protein complex comprised of a proline-rich enamel protein, SCPPPQ1, and laminin 332 (LAM332) to enhance the JE attachment. Using quartz crystal microbalance with dissipation monitoring (QCM-D), we showed that SCPPPQ1 and LAM332 interacted and assembled into a protein complex with high-affinity adsorption of 5.9e(-8) [M] for hydroxyapatite (HA), the main component of the mineralized tooth surfaces. We then designed a unique shear device to study the adhesion strength of the oral epithelial cells to HA. The SCPPPQ1/LAM332 complex resulted in a twofold enhancement in adhesion strength of the cells to HA compared to LAM332 (from 31 dyn/cm(2) to 63 dyn/cm(2)). In addition, using a modified wound-healing assay, we showed that gingival epithelial cells demonstrated a significantly high migration rate of 2.7 +/- 0.24 mu m/min over SCPPPQ1/LAM332-coated surfaces. Our collective data show that this protein complex has the potential to be further developed in designing a bioadhesive to enhance the JE attachment and protect the underlying connective tissue from bacterial invasion. However, its efficacy for wound healing requires further testing in vivo. Statement of significance This work is the first functional study towards understanding the combined role of the enamel protein SCPPPQ1 and laminin 332 (LAM332) in the epithelial attachment of the gum, the junctional epithelium (JE), to the tooth hydroxyapatite surfaces. Such studies are essential for developing therapeutic approaches to restore the integrity of the JE in the destructive form of gum infection. We have developed a model system that provided the first evidence of the strong interaction between SCPPPQ1 and LAM332 on hydroxyapatite surfaces that favored protein adsorption and subsequently oral epithelial cell attachment and migration. Our collective data strongly suggested using the SCPPPQ1/LAM332 complex to accelerate the reestablishment of the JE after surgical gum removal to facilitate gum regeneration. (c) 2022 The Authors. Published by Elsevier Ltd on behalf of Acta Materialia Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)

Introduction Periodontitis is characterized by the destruction of tooth-supporting tissues including the alveolar bone. Barrier membranes are used in dentistry for tissue regenerative therapy. Nevertheless, conventional membranes have issues related to membrane stability and direct induction of bone mineralization. Amelotin (AMTN), an enamel matrix protein, regulates hydroxyapatite crystal nucleation and growth. To apply an AMTN membrane in clinical practice, we investigated the mineralizing and adhesive effects of recombinant human (rh) AMTN in vitro using a collagen-based system. Methods Collagen hydrogel incorporated with rhAMTN (AMTN gel) and rhAMTN-coated dentin slices were prepared. AMTN gel was then applied on a commercial membrane (AMTN membrane). Samples were incubated for up to 24 h in mineralization buffer, and the structures were observed. The peak adhesive tensile strength between the dentin and AMTN membrane was measured. Using an enzyme-linked immunosorbent assay, the release kinetics of rhAMTN from the membrane were investigated. Results The AMTN gel resulted in the formation of hydroxyapatite deposits both onto and within the collagen matrix. Furthermore, coating the dentin surface with rhAMTN promoted the precipitation of mineral deposits on the surface. Interestingly, site-specific mineralization was observed in the AMTN membrane. Only 1% of rhAMTN was released from the membrane. Hence, the AMTN membrane adhered to the dentin surface with more than twofold greater tensile strength than that detected for a rhAMTN-free barrier membrane. Conclusions RhAMTN can accelerate mineralization and adhesion in collagen-based systems. Furthermore, the AMTN membrane could inform the optimal design of calcified tissue regenerative materials.